Microfluidic Selection of Aptamers for Biological Purification Applications Principal Investigator: Qiao Lin, Columbia University Aptamers are oligonucleotides that bind to small molecules or proteins. Aptamers are isolated through an in vitro selection and amplification procedure called systematic evolution of ligands by exponential enrichment (SELEX). The procedure is based on an affinity selection, followed by amplification of nucleic acids that are binders from large random libraries. Aptamers can be developed for an extremely broad spectrum of analytes with high affinity, can possess well controlled target selectivity, and can be synthesized to bind targets with predefined characteristics. In particular, aptamer binding in general exhibits strong temperature dependence; thus, aptamers may specifically bind target analytes at a predefined temperature and reversibly decouple from the targets at a modestly different, yet also predefined, temperature. This property is attractive to biomedical applications such as affinity purification, as it can enable specific purification with thermally activated release and isocratic elution of analytes. Conventional SELEX instruments have enabled great progress in aptamers, but their use is generally labor-intensive and time-consuming. These limitations can be addressed by leveraging microfluidic technology. We propose to pursue proof-of-concept demonstration of a microfluidic SELEX system that integrates all steps of the SELEX method to allow automated development of aptamers with predefined temperature-dependent binding characteristics for applications to affinity purification of analytes. Our specific aims include: (1) developing a bead-based polymerase chain reaction (PCR) technique in a microchannel to establish its applicability to microfluidic SELEX; (2) integrating microchip DNA selection and amplification to create a prototype microfluidic SELEX system; and (3) characterize and validate the prototype system by using it to select aptamers against well established proteins. The proposed research is relevant to public health because it will provide a platform for selecting aptamers for affinity purification of analytes such as proteins, small molecules and cells involved in biomedical research on diseases. In addition, this system could also be used to develop aptamers for applications to target validation, drug discovery, diagnostics, and therapy. PUBLIC HEALTH RELEVANCE (provided by applicant): Microfluidic Selection of Aptamers for Biological Purification Applications Principal Investigator: Qiao Lin, Columbia University This research will pursue proof-of-concept demonstration of a microfluidic system that integrates all steps of the method of systematic evolution of ligands by exponential enrichment (SELEX). The system will allow automated development of aptamers with predefined temperature-dependent binding characteristics for analyte purification applications. It is relevant to public health because it will provide a platform for selecting aptamers for affinity purification of analytes such as proteins, small molecules and cells involved in biomedical research on diseases. In addition, this system could also be used to develop aptamers for applications to target validation, drug discovery, diagnostics, and therapy.